HIGH THROUGH-PUT AUTOMATED DNA SEQUENCING

高通量自动化 DNA 测序

• 批准号: 6054035
• 负责人: Judith L CAMPBELL
• 金额: $ 16.06万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-04-01 至 2001-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6054035
• 关键词: biomedical equipment purchase; high throughput technology; nucleic acid sequence

We request funding for instrumentation to update a successful central DNA sequencing core facility at the California Institute of Technology. The Core Facility has developed over the last five years into an indispensable contributor to the ongoing research projects of Biology, Chemistry, Geology, and Environmental Engineering Divisions. Currently we are producing about 2500 sequences a month for walk-in users. In addition, the Core Facility has carried out two production scale sequencing projects coordinated by the Genome Resource Center at Caltech, completing the sequence of an entire micro-organism, Pyrobaculum aerophilum, and a large region of human chromosome 16, in addition to many BAC clones. All labs at Caltech now rely on automated, fluorescent-based sequencing. In order to continue the quality of service we have achieved in the face of increasing demand, we are requesting three types of equipment: an ABI Prism 3700 DNA Analysis system, new PCR machines, and several small computers. We have chosen the ABI Prism 3700, which is based on a novel capillary electrophoresis system rather than slab gels, because of its promise of the highest through-put, with the least labor at the lowest cost and with the highest quality of any such instrument currently available. The PCR machines will be needed to exploit the capacity of the ABI3700. The computers are needed to interface the new equipment with the existing Sequence Analysis Facility which distributes the sequence to the users. Twenty-eight major users provide abstracts of the work they will do. There will continue to be standard sequencing of single cDNAs, PCR products, site-directed mutants, etc. This will provide service to projects covering, among others, genes involved in RNA splicing, molecular control of neural crest and/or muscle development, in controlling body segment development in Drosophila, in cell cycle regulation in Xenopus and yeast, in cell fate and morphogenesis in several organisms, genes involved in floral development in Arabidopsis. Availability of the enormous capacity of the 3700 will, we hope, also drive new approaches in the laboratories here. For instance, several labs (Anderson, Wold, Meyerowitz) are developing strategies to identify cDNA markers based on differential gene expression for various specific cell types and individual stages of development. Another project proposes to screen age-related mutations in human mitochondria. Another dimension is sequencing entire microorganisms and panning for organisms with interesting known pathways. These latter projects could involve many thousands of runs per month in themselves. Thus, we hope to maintain our ability to function both as a walk-in facility and a high-tech production scale facility, as we have done in the past.

我们要求提供仪器的资金，以更新加州理工学院成功的中央DNA测序核心设施。 在过去的五年里，核心设施已经发展成为生物学，化学，地质学和环境工程部门正在进行的研究项目不可或缺的贡献者。 目前，我们正在生产约2500序列一个月的步行用户。 此外，核心设施还在加州理工学院基因组资源中心的协调下进行了两个生产规模的测序项目，完成了整个微生物Pyrobaculum aerophilum和人类16号染色体的大部分区域以及许多BAC克隆的测序。 加州理工学院的所有实验室现在都依赖于自动化的荧光测序。 为了继续保持我们在不断增长的需求面前所取得的服务质量，我们需要三种类型的设备：ABI Prism 3700 DNA分析系统，新的PCR仪和几台小型计算机。 我们选择了ABI Prism 3700，它是基于一种新型的毛细管电泳系统，而不是平板凝胶，因为它的承诺，最高的吞吐量，最少的劳动力，最低的成本和最高的质量的任何此类仪器目前可用。 需要PCR机器来利用ABI 3700的能力。 需要计算机将新设备与现有的序列分析设备连接起来，后者将序列分发给用户。28个主要用户提供了他们将要做的工作的摘要。 将继续对单个cDNA、PCR产物、定点突变体等进行标准测序。这将为涉及RNA剪接、神经嵴和/或肌肉发育的分子控制、果蝇体节发育的控制、非洲爪蟾和酵母的细胞周期调节、几种生物体的细胞命运和形态发生、拟南芥花发育相关基因。我们希望，3700的巨大容量的可用性也将推动这里实验室的新方法。 例如，几个实验室（安德森，沃尔德，迈耶罗维茨）正在开发的战略，以确定cDNA标记的基础上差异基因表达的各种特定的细胞类型和个别阶段的发展。 另一个项目提出在人类线粒体中筛选与年龄相关的突变。 另一个方面是对整个微生物进行测序，并通过有趣的已知途径筛选生物体。 后者的项目本身可能涉及每月数千次运行。因此，我们希望保持我们的能力，既作为一个步入式设施和高科技生产规模的设施，因为我们已经做了过去。